Histone deacetylase activity in conjunction with E2F-1 and p53 regulates Apaf-1 expression in 661W cells and the retina.
Apaf-1 and the cysteine proteases known as caspases are genes central to the intrinsic apoptotic pathway in the retina. Previously, we have shown that histone deacetylase (HDAC) activity regulates Apaf-1 expression in the retina. In this study, we unravel the detailed molecular mechanism of HDAC-mediated regulation of Apaf-1 initially by use of a cell line (661W), which expresses some cone-specific genes and then by means of an ex vivo retinal explant system. Inhibition of HDAC activity by trichostatin A (TSA) up-regulates Apaf-1 expression, which precedes the induction of apoptosis. Furthermore, by a bioinformatics approach, we identify E2F-1 and p53 binding sites on the mouse Apaf-1 promoter and show by chromatin immunoprecipitation assays that these sites are occupied in vitro and that treatment with TSA results in increased binding of E2F-1 and p53 to the Apaf-1 promoter. By performing siRNA to these transcription factors, we illustrate that they govern Apaf-1 expression levels in vitro. Finally, in a retinal explant system, we show that similar to our 661W results, E2F-1 and p53 are up-regulated after inhibition of HDAC activity in the retina. This correlates with our previous observation in the explant system that Apaf-1 expression increases significantly and leads to an induction of apoptosis after inhibition of HDAC activity. Overall, we propose a role for HDAC activity, E2F-1, and p53 in the regulation of Apaf-1 expression in 661W cells; initial data also indicate a regulatory role in the retina.
Wallace
Deborah M
DM
Cell Development and Disease Laboratory, Department of Biochemistry, Biosciences Institute, University College, Cork, Ireland.
Cotter
Thomas G
TG
D000107:Acetylation / Q000187:drug effects; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D053477:Apoptotic Protease-Activating Factor 1 / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002087:Butyrates / Q000494:pharmacology; D002460:Cell Line; D050687:E2F1 Transcription Factor / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D056572:Histone Deacetylase Inhibitors; D006655:Histone Deacetylases / Q000378:metabolism*; D006877:Hydroxamic Acids / Q000494:pharmacology; D066298:In Vitro Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012160:Retina / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D015854:Up-Regulation
C506593:Apaf1 protein, mouse; D053477:Apoptotic Protease-Activating Factor 1; D002087:Butyrates; D050687:E2F1 Transcription Factor; C493754:E2f1 protein, mouse; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; C012589:trichostatin A; D006655:Histone Deacetylases